Vaccines
The production of a protective immune response against any given infection agent in vertebrates depends initially on the provision of the appropriate stimulus to the host's immune system. The infectious organism itself typically provides numerous stipulatory compounds, or antigens, by the very nature of its cell membrane composition, or by the metabolic products it releases in the host's body. These substances, usually larger molecules such as proteins, lipopolysaccharides or glycoproteins, are recognized by the immune system as foreign, and provoke one or more different types of reaction from the host in an effort to remove or disable the invading organism. The antigen may cause production of sensitized lymphocytes (T-cells) which provide a cell-mediated immunity. Alternatively, an antigen may stimulate the synthesis and release of free antibody into the blood and other bodily fluids (humoral immunity). The development of the body's protective immune response depends upon achieving a threshold level of stimulation of one or both of these systems.
A temporary immunity against infection can in many cases be provided by giving an individual preformed antibodies from another individual of the same or different species. This is known as passive immunity. One example of such immunity is the protection afforded to a fetus and newborn by placental transfer of maternal antibodies, as well as transfer of antibodies through milk. Another example is the pooled adult gamma globulin frequently used to prevent or modify the effects of exposure to measles, chicken pox, hepatitis, smallpox and tetanus. These acquired antibodies, however, are gradually utilized by interaction with the antigen or catabolized by the body, and thus the protection is eventually lost.
A more permanent form of protection is afforded by active immunization. Vaccination confers an active protective immunity by employing a harmless or nonvirulent form of the antigen (e.g., a killed or genetically altered bacterium, or an isolated glycoprotein from the cell wall) as a primary stimulus to the immune system. This provokes a rather slow response in antibody production which peaks and falls off. However, the body has been alerted to the existence of the antigen, and the next time exposure occurs, presumably with the live, virulent organism, a secondary response, with a much more rapid and abundant production of antibodies is observed. This secondary response will typically be sufficient to prevent the microorganism from establishing itself sufficiently to be able to cause a full-blown infection.
A vaccine may take a variety of forms, some of which are more effective than others in conferring the protective effect desired. Historically many vaccines have been prepared by killing or inactivating the microorganism causing the disease of interest, and using the killed cells as the active immunogenic agent. Vaccines of this type have been used against typhoid, cholera and poliomyelitis (Salk vaccine). The problem with this type of vaccine is that the treatment required to kill the microorganisms, such as formaldehyde, may frequently alter or destroy the microbe's useful antigens, and thus poor or incomplete immunity may be obtained.
An alternate form of vaccine is that which employs attenuated microorganisms. An attenuated microorganism is one which is still living, and capable of multiplying in the body after administration, but which has either been modified in some way to render it avirulent, or else is a strain which is virulent in other microorganisms, but avirulent in man. The advantage obtained by using an attenuated organism is that the attenuation usually does not affect its antigenicity, thereby providing a more efficient stimulus to the immune system. Attenuated vaccines for measles, rubella and poliomyelitis (Sabin vaccine) have achieved widespread use. Attenuation is typically achieved by altering the growth conditions of the microorganism, or, more recently, genetically modifying the organism's virulence. Although generally more effective than killed vaccines, however, there is danger involved in the possibility of the live organisms reverting to virulence, thereby causing disease symptoms.
Because of the problems associated with whole organism vaccines, it has become more common in recent years to employ individual protective antigens as the active agent in vaccine compositions. Although isolation and identification of the particular antigens of an organism which do stimulate a protective response is not always simple, once identified, this method provides an effective alternative to the whole organism vaccines, without the attendant disadvantages.
Examples of types of antigens which are typically useful for this purpose are purified cell or capsid components, such as bacterial toxins (which must be detoxified to yield a toxoid), bacterial or viral toxin subunits, cell wall polysaccharides, or capsid glycoproteins. These components are often highly effective in producing immunity, but difficulties arise in the actual purification. It is critical that the antigen of interest be isolated more or less completely from extraneous cellular materials, the presence of which can frequently cause an adverse immunologic or metabolic response concurrent with the desired immunity producing reaction. The purification processes required are frequently complex, tedious and prohibitively expensive and the results not always completely predictable. This problem can in some case be avoided by synthetic preparation of small peptide sequences which correspond to epitopes on a microbial antigen. There are, in some circumstances, drawbacks to this technique in that, although the amino acid sequence corresponds to the natural epitope sequence, the conformation may not be sufficiently similar to that of the parent antigen to elicit the appropriate immune response.
Many of the disadvantages attendant upon these aforementioned vaccine types may be avoided by the use of recombinant DNA technology. The possibility of cloning genes which code for all or part of an important microbial antigen provides a relatively economical and convenient source of larger amounts of the necessary immunogenic material, substantially free of contaminating proteinaceous or genetic material. In brief, this method of producing purified antigens involves the insertion of the specific DNA sequence coding for the antigen into a DNA vector to form a recombinant DNA molecule which can be replicated by a host cell. There is now an abundance of methods by which recombinant DNA molecules may be prepared. One of the better known techniques is that described by Cohen and Boyer in U.S. Pat. No. 4,237,224, the teachings of which are herein incorporated by reference. In this method recombinant plasmids are produced using restriction enzyme and ligation; the resulting plasmids are then placed into a unicellular host cell which is thereby transformed, and begins replication of the foreign DNA. Another method, utilizing bacteriophage vectors, is described in U.S. Pat. No. 4,304,863, also incorporated herein by reference. The cloning method provides the greatest potential for making convenient, economical sources for vaccine preparation readily available; it is not without difficulties, however, in that the appropriate gene for an antigen known to be immunogenic must first be isolated and sequenced, inserted into a plasmid in such a way as to insure proper replication, transcription and translation, and then inserted into a compatible host cell.
The potential for failure of the expression of the desired gene product is tremendous if any one of transcription of the gene, translation of the RNA, or post-translational processing and compartmentalization of the polypeptide is not performed correctly. For a cloned insert to be transcribed, it is necessary that there is present a promoter which the host rNA polymerase can recognize. Proper translation requires that the RNA has a ribosome binding site. Post-translational modification of proteins often involves cleaving of a signal sequence which functions by directing the protein out through the cell membrane. Degradation of the foreign protein produced by the host microorganism may also occur if the configuration or amino acid sequence does not protect them from the action of the host's intracellular proteases. Thus, although conceivably the ideal way of ultimately constructing a Neisseria vaccine, these techniqes have not previously been applied to vaccine preparation for gonorrhea.